Adult bone marrow (BM)-derived hematopoietic stem cells (HSC) are defined by their ability to self renew while functionally repopulating the cells of the blood and lymph for the life of an individual. See, Müller-Sieburg, C. (ed.) Hematopoietic stem cells: animal models and human transplantation (Springer-Verlag, New York, 1992). These abilities make HSC clinically useful in therapeutic BM transplantation for a variety of BM failure states including the hematological malignancies leukemia and lymphoma. HSC can be highly enriched and quantified by known methods. See, e.g., Harrison et al. Exp Hematol 21, 206-19 (1993). Like other tissue-derived stem cells, HSC are thought to retain a high capacity for “plasticity” that would allow for the potential contribution of regenerative progenitors to non-hematopoietic tissues following injury or stress. Goodell et al., Ann N Y Acad Sci. 938, 208-18; discussion 218-20 (2001); Krause et al., Cell 105, 369-77 (2001).
Indeed, following full and durable reconstitution of a lethally irradiated mouse with a single BM-derived HSC, donor cells were identified in multiple tissues such as the brain, heart, skeletal muscle, liver, and endothelial cells. Krause et al., supra. Although the experimental design of that study yielded low levels (<5%) of donor contribution to non-hematopoietic tissues, the results suggest the possibility of functional regeneration of multiple tissues by HSC-derived progenitors. In other transplant models hematopoietic progenitors have been shown to repopulate hepatocytes in the parenchymal liver to restore liver function following chemically induced injury (Petersen et al., Science 284, 1168-70. (1999); Lagasse et al., Nat Med 6, 1229-34 (2000)), and to regenerate myocardium to improve cardiac function following infarction (Orlic et al., Nature 410, 701-5 (2001)).
Diabetic retinopathy and retinopathy of prematurity are among the leading causes of vision impairment throughout the world. Retinal neovascularization is thought to occur in response to an hypoxic insult which leads to changes in the existing vasculature and compensatory, albeit pathologic, new capillary growth. Grant et al., Diabetes 35, 416-20 (1986); Limb et al., Br J Ophthalmol 80, 168-73 (1996). Postnatal neovascularization has been attributed to angiogenesis, a process characterized by sprouting of new capillaries from pre-existing blood vessels. Folkman and Shing, J Biol Chem 267, 10931-4 (1992). Several studies have shown that endothelial progenitor cells (EPC) capable of contributing to in vitro capillary formation can be derived from BM cells. Asahara et al., Science 275, 964-7 (1997); Gehling et al., Blood 95, 3106-12 (2000); Bhattacharya et al., Blood 95, 581-5 (2000); Lin et al., J Clin Invest 105, 71-7 (2000). Pro-angiogenic growth factors such as vascular endothelial growth factor (VEGF, See, e.g., Asahara et al., Embo J 18, 3964-72 (1999); Kalka et al., Circ. Res. 86, 1198-202 (2000)), and granulocyte/macrophage colony stimulating factor (GM-CSF) (see, e.g., Takahashi et al., Nat Med 5, 434-8 (1999)) increase circulating levels of EPC in the adult and promote new blood vessel formation. Recently, it was demonstrated that hydroxymethylglutaryl (HMG)-CoA reductase inhibitors potently augment EPC differentiation by a mechanism involving the angiogenic protein kinase Akt. Dimmeler, et al., J Clin Invest 108, 391-7 (2001). Studies also support the contribution by EPC to blood vessel formation in the adult (Asahara et al., Embo J 18, 3964-72 (1999); Kalka et al., supra; Crosby et al., Circ Res 87, 728-30 (2000); Murohara et al., J Clin Invest 105, 1527-36 (2000)), and in cardiac reperfusion post ischemia (Kocher et al., Nat Med 7, 430-6 (2001); Kawamoto et al., Circulation 103, 634-7 (2001)). However, as these studies were based on short-term transplant and acute injury models, it is not clear whether the cell giving rise to EPCs is the long-term repopulating HSC or other progenitors such as the mesenchymal stem cell.
Within the developing embryo, pluripotent progenitors are generated that are capable of contributing to the formation of blood and blood vessels, a process called hemangiogenesis. Choi, K., Biochem Cell Biol 76, 947-56 (1998); Takakura, et al., Cell 102, 199-209 (2000). These pluripotent stem cells are termed hemangioblasts. Hemangioblasts can also be produced from embryonic stem cells during in vitro differentiation in response to vascular endothelial growth factor. Choi, supra. Heretofore, however, definitive evidence for the existence of the hemangioblast within the adult BM, and in particular for a functional role of such BM-derived cells in new blood vessel formation was lacking.